Work in our laboratory is aimed at understanding the molecular events governing development of the skeleton, as skeletal defects reflect 25% of all human congenital malformations. We are engaged in research to bridge the gap of understanding between genetic mutations in MSX2 and FGFR2 and the resulting defects in human craniofacial and limb skeletal development. In addition to our studies to analyze the function of these human disease genes, we have initiated a molecular search for new genes that can serve as rate limiting regulators for early events in chondrogenesis. Signal transduction events initiated by bone morphogenetic proteins (BMP's) and fibroblast growth factors (FGF's) are important in modulating cell proliferation, migration, differentiation, apoptosis and tissue patterning during the development of several organ systems including the skeleton. We are testing a series of hypotheses regarding the function of the homeodomain transcription factor Msx2 as a point of integration of signals from the BMP and FGF mediated pathways. We have examined the expression profiles of BMP4, FGF8, Msx1 and Msx2 in the developing mouse embryo during organogenesis. The deduced dynamic, temporally and spatially restricted localization patterns of these molecules have served as a foundation for the construction of new hypotheses of the molecular mechanisms of skeletal ontogeny. Our experiments have shown that Msx2 functions in the apoptotic elimination of cranial neural crest cells where it is normally expressed in rhombomeres 3 and 5 of the embryonic hindbrain. Gain of function, ectopic expression of Msx2 in rhombomeres 4 and 6 induced apoptosis and eliminated neural crest cell emigration from these structures in vivo and in tissue culture. These studies confirmed when and where crest cells emigrate into the first branchial arch. We have identified morphoregulatory compartments in the developing mandible by implanting growth factor soaked beads and monitoring subsequent gene expression and cell type differentiation adjacent to the source of signalling. BMP4 induced Msx2 expression and caused precocious Meckel's cartilage expansion or bifurcation, or had no effect on cartilage, depending on the topographical position of the bead implantation relative to these putative tissue compartments. To identify other signaling pathways that contribute to the regulation of chondrogenesis, we have designed and characterized a specialized tissue culture system for examining the effects of compressive force on chondrogenesis. We have found that static compressive force can stimulate the differentiation of chondrocytes from mesenchymal cells by regulating the expression of the transcription factor Sox9 and the cytokine IL1-beta. These and other projects are in progress, expanding our understanding of the molecular mechanisms of chondrogenesis. The synergy of these projects will contribute to deciphering the complex interactions of signal transduction pathways stimulated by FGF's, BMP's, and cell adhesion as they impact on normal skeletal development and common skeletal malformations.